2003 年 72 巻 2 号 p. 79-86
To evaluate the N-glycosylation property of an insect cell line, NISES-AnPe-428 (AnPe), which was derived from embryos of the Chinese oak silkworm, Antheraea pernyi, and used in a newly developed baculovirus expression vector (BEV) system, N-glycans of purified prothoracicotropic hormone (PTTH) produced by AnPe cells infected with a recombinant A. pernyi nucleopolyhedrovirus (AnpeNPV), were digested with glycoamidase A (from sweet almond), reductively aminated with 2-aminopyridine, and identified by two-dimensional high performance liquid chromatography (2D-HPLC) mapping technique. For comparison, N-glycans of PTTH produced by Spodoptera frugiperda cells (Sf9) infected with a recombinant Autographa californica NPV (AcNPV) were also analyzed similarly. Consequently, similarities and differences in the N-glycan structure composition between AnPe and Sf9 were revealed. N-glycans from AnPe contained 41.8% high mannose type, 32.8% paucimannosidic type, 12.5% monoantennary type, and 1.3% biantennary, complex type with terminal N-acetylglucosamine (GlcNAc) residues, while those from Sf9 contained 46.5% high mannose type, 29.8% paucimannosidic type, 7.7% monoantennary type, and no detectable biantennary complex type. In addition, one of the monoantennary type N-glycans with a terminal GlcNAc residue identified in AnPe was not detected in Sf9. The proportion of N-glycans with an α1, 6-linked fucose residue at the innermost core GlcNAc residue was 41.4% in AnPe and 23.4% in Sf9, respectively. In both cell lines, mammalian-like complex N-glycans with terminal galactose and/or sialic acid residues were not identified. These results indicated that a reaction catalyzed by GlcNAc transferase II (GnTII), a key enzyme to form biantennary, complex N-glycans, occurs more efficiently in AnPe than in Sf9, suggesting that the proportion of complex N-glycans in AnPe cells may be largely enhanced by inhibiting the N-acetylglucosaminidase activity and/or supplying sufficient amounts of enzymes and substrates required for the sugar chain extension in the Golgi.
